Project Summary/Abstract Background: The intestinal epithelium represents one of the largest exposed surface areas of the body and must perform multiple functions related to digestion, infectious surveillance, and metabolism. Helping to coordinate these activities are enteroendocrine cells (EECs), specialized intestinal epithelial cells expressing cellular machinery for chemical and mechanical sensing and for host communication via the ?Gut-Brain Axis.? Liver receptor homolog-1 (LRH-1, NR5A2) is a nuclear receptor expressed in the intestinal epithelium. We have shown LRH- 1 is critical for the production of these unique cells and that loss of LRH-1 leads to a remarkable visceral hyposensitivity. In this application, we propose to elucidate both the mechanism(s) by which LRH-1 promotes the differentiation of EECs and the functional consequences of their loss on gut-brain communication. Approach: We will use a combination of genetic mouse models and intestinal organoid technology to investigate LRH-1- dependent EEC differentiation. Using a single cell sequencing approach, we will identify EEC progenitor population shifts following loss of LRH-1. Leveraging an EEC progenitor fluorescent marker, we will observe in real time the effects of LRH-1 knockout on EEC cell differentiation and to delineate the LRH-1 DNA binding targets. Our molecular studies will be complemented by an evaluation of the functional consequences of LRH- 1-dependent EEC loss. Finally, use of detailed anatomic studies in our acute LRH-1 knockout models will enable us to explore the effects of rapid EEC disruption on mucosal nerve fibers, a key component of the gut- brain axis. Goals: This proposal is designed to expand upon a key finding uncovered during my K08 investigation and to fully establish my independent research program. These experiments will provide important data for submission of a successful R01 proposal built at the extraordinary confluence of intestinal epithelial physiology, nuclear receptor biology, and neuronal signaling.